Earlier studies from several laboratories on the changes in the ultrastructure and enzymic activities of mitochondria from senescent animals have led to the hypothesis that the mitochondrial membrane becomes fragile in aging. We propose to analyze the biochemical mechanisms that underlie mitochondrial membrane degradation and subsequently determine if these reactions are affected in aging. The methods for measuring mitochondrial concentration in cultured hepatocytes will be devised, and the changes in mitochondrial concentration determined at different times after producing controlled damage to the membrane by the use of respiratory inhibitors. The fate of a single inner mitochondrial membrane protein will be followed using a specific labeling technique. The protein and partially degraded polypeptides from it will be obtained by immunoprecipitation. The relative roles of lysosomes and cytosolic proteases in the breakdown of the labeled membrane protein will be explored. In subsequent phases of the program, we will examine the regulation of the degradation and what changes occur in aging.